Capillary electrophoresis apparatus and method

ABSTRACT

An electrophoresis apparatus is provided in which negative effects caused by abnormalities in a current-carrying path in an electrophoresis apparatus can be avoided or reduced. The current flowing in the current-carrying path during electrophoresis can be measured to detect the state of a separating medium, and the application of a voltage to the current-carrying path can be interrupted.

FIELD

[0001] The present teachings relate to a capillary electrophoresisapparatus for separating and analyzing samples, such as nucleic acids orproteins.

BACKGROUND

[0002] A known capillary electrophoresis apparatus is the Prism 3100, byApplied Biosystems. This apparatus can detect an electric currentflowing between an electrode in a cathode-side buffer solution and ahigh voltage source, and an electric current flowing between anelectrode in an anode-side buffer solution and ground.

SUMMARY

[0003] The present teachings provide a capillary electrophoresisapparatus that can avoid or reduce problems caused by defects in acurrent-carrying path.

[0004] The present teachings relate to an apparatus and method formeasuring an electric current flowing in a current-carrying path duringelectrophoresis to detect the state of the current-carrying path. Anabnormality, such as the growth of air bubbles, can be detected based onchanges in the electric current in the current-carrying path. Acountermeasure, such as the termination of electrophoresis, can beundertaken upon detecting an abnormality. As a result, adverse effectssuch as electric discharge that can be caused by continuingelectrophoresis after an abnormality has occurred, can be avoided.

[0005] The present teachings relate to an apparatus and method that canmeasure an electric current flowing in a current-carrying path when apredetermined voltage is applied during electrophoresis for detectingthe state of the current-carrying path. A determination can be madewhether the current-carrying path is suitable for electrophoresis basedon the detected value of the electric current when a voltage that wouldnot cause adverse effects such as electric discharge is applied. As aresult, the state of a separating medium can be easily determined, andadverse effects, such as electric discharge, can be avoided even in anabnormal situation, such as when the capillaries are not filled with aseparating medium.

[0006] Additional features and advantages of various embodiments will beset forth in part in the description that follows, and in part will beapparent from the description, or may be learned by practice of variousembodiments. The objectives and other advantages of various embodimentswill be realized and attained by means of the elements and combinationsparticularly pointed out in the description herein and appended claims.

BRIEF DESCRIPTION OF THE DRAWINGS

[0007]FIG. 1 schematically shows an electrophoresis apparatus accordingto various embodiments;

[0008]FIG. 2 is a circuit diagram of a high-voltage power supply thatcan constitute a voltage control device of the electrophoresis apparatusaccording to various embodiments;

[0009]FIG. 3 shows a flowchart from the start of an analysis to the endaccording to various embodiments;

[0010]FIG. 4 shows a flowchart for detecting abnormalities in anelectrophoresis path according to various embodiments;

[0011]FIG. 5 shows a method of detecting and comparing electric currentfluctuations according to various embodiments; and

[0012]FIG. 6 graphically illustrates electric current changes andfluctuations according to various embodiments.

[0013] It is to be understood that both the foregoing generaldescription and the following detailed description are exemplary andexplanatory only, and are intended to provide an explanation of variousembodiments of the present teachings.

DETAILED DESCRIPTION OF THE VARIOUS EMBODIMENTS

[0014]FIG. 1 schematically shows an electrophoresis apparatus accordingto various embodiments. The structure of the electrophoresis apparatuswill be described by referring to FIG. 1.

[0015] According to various embodiments, the apparatus can include adetection cell 116 that can enable optical detection of samples, athermostatic oven 118 that can maintain the capillaries at a constanttemperature, a transporter 125 that can transport various containers toa capillary cathode end, a high-voltage power supply 119 that can applya high voltage to the capillaries, a first ammeter 120 that can detect acurrent supplied by the high-voltage power supply 119, a second ammeter112 that can detect a current flowing in an anode electrode, a capillaryarray 117 that can include one or more capillaries, and a pump device102 that can inject a separation medium into the capillaries.

[0016] According to various embodiments, the capillary array 117 can bea replacable member including 96 capillaries. The capillary array 117can include a block 129, a detection cell 116, and a capillary head.When a method of analysis is changed, the capillary array can bereplaced and the lengths of the capillaries can be varied. When there isdamage to or deterioration of the quality of the capillaries, thecapillary array 117 can be replaced with a new one.

[0017] According to various embodiments, each capillary can be made offused silica, for example, and can have an internal diameter rangingfrom 10 to several hundred microns, and can have an external diameter ofat least 100 microns. The surface of the capillaries can be coated withpolyimide, for example, to increase strength, except for alight-illuminable portion where the coating is not applied or has beenremoved. The light-illuminable portions can allow light that has beenemitted by the sample migrating within the interior of the capillaries,for example, to be emitted to the exterior of the capillaries. Theinterior of a capillary can be filled with a separating medium to impartdifferences in migration speed during electrophoresis. Both a fluid anda non-fluid separating medium can be used. For example, a fluidseparation medium can be implemented as the separation medium accordingto various embodiments.

[0018] According to various embodiments, the capillary array can,instead of including a plurality of separate capillary tubes, include asubstrate such as a glass or plastic substrate having a plurality ofcapillary channels formed therein. The capillary channels can be formedby, for example, grooving techniques, etching techniques, maskingtechniques, molding techniques, chemical vapor deposition, and othermanufacturing methods, or a combination thereof. Herein, the term“capillary electrophoresis” includes electrophoresis using such asubstrate. When so provided, the capillary channels of the substrate caneach have a respective illuminable portion that is separated, by thesubstrate material, from the illuminable portions of adjacent capillarychannels.

[0019] According to various embodiments, the detection cell 116 can be acomponent for acquiring sample-dependent information. When the detectioncell 116 is illuminated with excitation light, it can emit light havingwavelengths that are dependent on the constituents of the sample. Thelight-illuminable portion and areas nearby to the 96 capillaries can bearranged and fixed to form an optically flat surface with a heighttolerance of a few microns. During electrophoresis, two substantiallycoaxial beams of laser light can be illuminated onto thelight-illuminable portions to the interior of the capillaries from bothsides of the detection cell 116, such that the light can be transmittedcontinuously through the light-illuminable portions to the interior ofthe capillaries. The laser light can cause the samples migrating withinthe capillaries to produce light (fluorescence having sample-dependentwavelengths) that can be emitted to the exterior of the capillariesthrough the light-illuminable portions. The emitted light can bedetected by an optical detector 115 and can be used to analyze thesamples.

[0020] According to various embodiments, capillary cathode end 127 ofthe capillaries can be fixed through a hollow metal electrode 126 sothat the tip of the capillary can protrude above the hollow electrode126 by about 0.5 mm. The hollow electrodes 126 of the individualcapillaries can be mounted on a block 129 as a single unit. The hollowelectrodes 126 can be connected to a high-voltage source 119 provided inthe apparatus main body, so that the hollow electrodes 126 can operateas cathode electrodes for applying voltages during electrophoresis orsample injection, for example.

[0021] According to various embodiments, filling end portions (arrangedopposite to the capillary cathode ends 127) of the capillaries can bebundled by a capillary head. The capillary head is a member that can beattached and detached in a sealed manner. The capillary head can beconnected to a block 107 in a pressure-tight and sealed manner. Thecapillaries can be filled with a fresh separation medium by a syringe106 through the filling end portions. The separation medium in thecapillaries can be replaced before each electrophoresis run to achieveimproved measurement performance.

[0022] According to various embodiments, a pump device 102 can include asyringe 106 and a mechanism for pressuring the syringe 106. The block107 can be a connecting portion for communicating the syringe 106, thecapillary array 117, an anode buffer container 110, and a separationmedium container 109. The pump device 102 system can be formed by anelectric motor 103, a linear actuator 132, and a transport member 136that can be adapted to have contact with the plunger of the syringe 106for transmitting force. When the electric motor 103 is turned in apositive direction, the transport member 136 can be pressed against theplunger of the syringe 106, thereby discharging the separation mediuminside the syringe 106 and injecting it into the capillaries. Thetransport member 136 can be provided with an electrically retractablehook 105. By turning the motor 103 in a reverse direction when the hook105 is hooked onto the plunger, the separation medium inside theseparation medium container 109 can be sucked out into the syringe 106.Positive rotation of the motor 103 is rotation of the motor in adirection in which the transport member 136 pushes the syringe, andreverse rotation is rotation in the opposite direction.

[0023] According to various embodiments, a check valve 108 can belocated between the separation medium container 109 and the block 107.The valve 108 can allow the flow of separation medium from theseparation medium container 109 to the block 107 while blocking the flowof the separation medium in the opposite direction. Thus, the checkvalve 108 can prevent the flow of separation medium back into theseparation medium container 109 when injecting it into the capillaryarray 117.

[0024] According to various embodiments, an electric valve 113 can beadapted to open or close the flow path between the block 107 and theanode buffer container 110. The electric valve 113 can close the flowpath at least when injecting separation medium into the capillaries, sothat the separation medium does not flow into the anode buffer container110. The valve can open the flow path when an electric current flowsbetween the anode and cathode, such as during electrophoresis.

[0025] According to various embodiments, an optical detection unit caninclude a light source 114 for illuminating the detection cell 116, andan optical detector 115 for detecting the light emitted from thedetection cell 116. When detecting samples separated by electrophoresisin the capillaries, the light-illuminable portion of the capillaries canbe illuminated by the light source 114, and the emission of light fromthe light-illuminable portion can be detected by the optical detector115.

[0026] According to various embodiments, the transporter 125 can includethree electric motors and linear actuators for transporting containersin three axial directions, namely vertical, horizontal, and depthdirections. A transport stage 130 of the transporter 125 can be capableof carrying at least one container. The transport stage 130 can beequipped with electrically-driven grips 131 by which individualcontainers can be held and released. The transporter 125 can transportthe buffer container 121, a washing container 122, a waste-liquidcontainer 123, and/or a sample container 124 to the cathode end of thecapillaries as necessary. Additional sample containers can be stored inpredetermined storage locations in the apparatus.

[0027] During operation, the main body 101 of the apparatus can beconnected with a computer 128 via communication cables. An operator canhave access to the data provided by the detectors in the apparatus bycontrolling the functions of the apparatus via the computer 128.

[0028]FIG. 2 shows a circuit diagram of a voltage-control deviceincluding a high-voltage power supply for the apparatus according tovarious embodiments. The voltage-control device will be described byreferring to FIGS. 1 and 2.

[0029] According to various embodiments, the voltage-control device caninclude a microprocessor 201, a controller 202, a high-voltage powersupply 203, a first ammeter 204, and a second ammeter 207. Thehigh-voltage power supply 203 can apply a voltage to a current-carryingpath under the control of the controller 202. The current-carrying pathcan include a hollow electrode 205, a buffer solution filled in thebuffer container 121, an electrophoresis path, a buffer solution filledin the anode buffer container 110, and an electrode (GND) 206. Theelectrophoresis path can include the separation medium filling thecapillaries, the block 107, and a connection pipe (fluid pipe connectingthe block 107 and the anode buffer container 110). According to variousembodiments, the first ammeter can be arranged between a high voltagepower supply and a hollow electrode, as shown in FIG. 2. According tovarious embodiments, the first ammeter can be arranged between a highvoltage power supply and ground, as shown in FIG. 1.

[0030] Referring to FIG. 2 and according to various embodiments, thehigh-voltage power supply 203 can be in electrical connection to thehollow electrode 205 via the first ammeter 204 and to the electrode(GND) 206 via the second ammeter 207. When about 5 to about 10 kilovoltsare applied across these electrodes, an electric field can be created ina direction from the hollow electrode 205 towards the electrode (GND)206. This electric field can cause a negatively charged sample, such asnucleic acids, to move from the capillary cathode end 127 to thedetection portion 116.

[0031] According to various embodiments, the first ammeter 204 candetect an electric current flowing from the high-voltage power supply203 to the hollow electrode 205, and can transmit the value of thecurrent to the microprocessor 201. The second ammeter 207 can detect anelectric current flowing from the electrode (GND) 206 to GND, and cantransmit the value of the current to the microprocessor 201. The valueof the electric current and its fluctuations can be checked by thesecond ammeter 207, as will be described later, because the secondammeter 207 can more directly indicate the value of the current flowingin the electrophoresis path.

[0032] In the event of electric leakage, for example, between the firstammeter 204 and the second ammeter 207, the value of current indicatedby the first ammeter 204 can include the value of the leaked electriccurrent, whereas the value indicated by the second ammeter 207 does notinclude the leakage component. Namely, the first ammeter 204 can detectthe net amount of the electric current flowing in the electrophoresispath. The interval between the first ammeter 204 and second ammeter 207includes media, such as the buffer and separation media, with relativelylarge resistance compared with metals, in addition to the connectingportions, such as the block and capillaries. Thus, the interval includesa part of the circuit of FIG. 2 that is liable to experience electricleakage.

[0033] According to various embodiments, the microprocessor 201 can readelectric current values from the first and second ammeters, 204 and 207,and can carry out calculations. The microprocessor 201 can then instructthe controller 202 to control the high-voltage power supply 203 into thestate of either a high-voltage application, a low-voltage application,or a forced voltage shutdown. The microprocessor 201 can be capable ofcommunicating with the externally located computer 128.

[0034] According to various embodiments, the various teachings can beused to cause a positively charged sample, such as a protein, to movefrom the capillary end 127 to the detection portion 116, by reversingthe cathode and anode ends.

[0035] According to various embodiments, preparations for conductingelectrophoresis will now be disclosed. Before initiating measurement,the operator can set up the following in the apparatus: the anode buffercontainer 121 holding the buffer solution; the washing container 122holding pure water for washing the capillaries; the waste-liquidcontainer 123 holding pure water into which the separation medium in thecapillaries is to be discharged; the separation medium container 109holding the separation medium; and the sample container 124 containingthe samples to be investigated.

[0036] According to various embodiments, the anode buffer container 110can be filled with a buffer to such a level that both the electrode(GND) 111 and the communicating tube are sufficiently submerged. Thecathode buffer container 121 can be filled with the buffer to such alevel that the hollow electrode 126 and the capillary cathode ends 127are sufficiently submerged. This can be done in light of the danger ofelectric discharge that could occur between the high electric-potentialcathode and other low electric-potential components upon the applicationof high voltage, if the amount of the buffer is insufficient or if themeasurement is initiated with the buffer container 121 empty. It is alsodesirable to have the buffer levels equal, so that the separation mediumin the capillaries is not moved by the pressure caused by heightdifferences.

[0037] According to various embodiments, the flow paths utilized duringelectrophoresis, or the flow paths utilized for transporting theseparation medium to those flow paths, can all be filled with separationmedium before the measurement starts. Normally, when the apparatus isused continuously, these flow paths can be filled with separationmedium. When the flow paths are refilled with separation medium afterthe capillary array is replaced or the flow paths are washed, theoperator can refill the flow paths with separation medium by operatingthe pump device 102 of the apparatus or by manually operating thesyringe 106.

[0038] The operator can then visually check to see if any air bubblesremain or any foreign substances are mixed in the flow paths. It ispossible that air bubbles or foreign substances can remain mixed due tooperational mistakes or improper checking. For example, micro-sized airbubbles can easily be missed by visual inspection. Such air bubbles orforeign substances in the flow paths could provide large resistance whenthe electrophoresis path is energized, possibly resulting in loweredanalysis accuracy due to deficient conduction, or damage to componentsdue to discharge. Even micro-sized air bubbles could be expanded byJoule heat during electrophoresis, eventually blocking the flow pathscausing electric discharge as disclosed above. Thus, the operator shouldpay attention to the condition of the flow paths before starting ananalysis. After setting the computer for a particular analysis to beinitiated, the operator can initiate the measurement. The analysis caninvolve applying a high voltage to the electrophoresis path.

[0039]FIG. 3 shows a flowchart that discloses the sequence from thestart of an analysis to the end of an analysis. The apparatus caninitiate an analysis in response to an instruction from the computer 128(301). First, the transporter, with which the apparatus is equipped, cantransport the waste-liquid container to the capillary cathode ends inpreparation for the injection of separation medium into the capillaries(302). Then, the pump device can inject a separation medium into thecapillaries (303). After a predetermined amount of separation medium isinjected, the transporter can transport the washing container to thecapillary cathode portion, where the capillary cathode ends can besubmerged and washed in the pure water in the washing container (304).The transporter can then transport the buffer container to the capillarycathode portion (305). Then, the condition of the current-carrying pathcan be confirmed (by confirming the value of the electric current uponapplication of a low voltage).

[0040] According to various embodiments, confirmation of the conditionof the current-carrying path can include a sequence including theapplication of a weak voltage (306), the checking of the current value(307), the displaying of errors (316), and a response by the operator(317). By this sequence, abnormalities such as, for example, a shortageof buffer solution, air bubbles remaining in the electrophoresis path,or the mixing in of dust can be identified. As a result, the danger ofcreating electric discharge in the electrophoresis path by theapplication of high voltage when there is a mistake made in thepre-preparation stage can be avoided.

[0041] According to various embodiments, in the step of applying a weakvoltage (306), a voltage can be applied that is lower than that appliedduring preliminary electrophoresis (308), sample introduction (311), andelectrophoresis (313). The magnitude of the voltage can be such that noelectric discharge is created, even if an abnormality, such as ashortage of buffer solution, air bubbles remaining in theelectrophoresis path, or the mixing of dust, has occurred. According tovarious embodiments, a voltage of about 1 kilovolt can be applied andthe value of the electric current can be confirmed about three secondslater. This can take place to obtain an accurate current valuecommensurate with the voltage, in consideration of the rise time of theelectric current. A wait time of about a few seconds can be sufficient.The value that is detected can then be read from the second ammeter 207on the anode side.

[0042] During the electric current check (307), the value of theelectric current detected by the weak voltage application (306) can becompared with a threshold. The threshold can be determined in view ofparameters that can influence the value of the electric current, such asthe kind of application, the specific length and number of thecapillaries, and the kind of separation medium used. Electric currentvalues that can be obtained under various use conditions can beexperimentally investigated, and a value that is about half or one-thirdof each of those current values can be used as the threshold. If thevalue of the electric current is smaller than the threshold, an abnormalstate or current-inhibiting factors, such as a shortage of buffersolution or the presence of air bubbles in the electrophoresis path, canbe assumed. As a result, an error can be presumed to exist in theelectrophoresis path. If such an abnormality is recognized, the voltagecan be shut down immediately to prevent damage to the components by wayof an electric discharge.

[0043] According to various embodiments, an error message can bedisplayed on the display of the control computer (316), and a responseby the operator can be demanded (317). Recognizing the error, theoperator can visually inspect the electrophoresis path for any airbubbles or a mixing of a foreign substance. If an abnormality isidentified, a countermeasure can be planned, and either the analysis canbe resumed or maintenance by service personnel can be requested. Forexample, if the cause of the abnormality turns out to be air bubbles orthe mixing of a foreign substance, the interior of the flow path can berefreshed by the pump device 102, or by manually operating the syringe106. The comparison of current values and the decision concerning anyabnormality can be carried out by software. The threshold used for thedecision concerning any abnormality can be stored in software.

[0044] According to various embodiments, confirmation of the conditionof the current-carrying path can be implemented prior to sampleintroduction (311) and electrophoresis (314). As a result, mistakes inthe initial preparation step can be avoided, such as a shortage of thebuffer volume resulting in the buffer solution failing to contact theelectrode, or the mixing of relatively large air bubbles in theelectrophoresis path causing a failure in establishing electricconduction.

[0045] According to various embodiments, if there is no abnormality inthe current-carrying path, a predetermined voltage can be applied and apreliminary migration can be initiated (308). The preliminary migrationcan be performed to optimize the condition of the separation medium inthe capillaries for analysis prior to the main analysis process fromsample introduction to electrophoresis. During preliminary migration, avoltage of about 10 kilovolts or more can be applied for about 2 or moreminutes. After preliminary migration, the capillary cathode ends can bewashed again in the washing container (309), and the sample container124 can be transported to the capillary cathode ends. When a voltage inthe order of about several kilovolts is applied to the capillary cathodeelectrode in the sample solution contained in the sample container 124,an electric field can be created between the sample solution and theanode electrode. This electric field can cause the samples in the samplesolution to be introduced into the capillaries (311).

[0046] After the sample is introduced, the capillary cathode ends can bewashed in the washing container (312), and then the buffer container canagain be transported to the capillary cathode end (313). A predeterminedvoltage can then be applied to start electrophoresis (314).

[0047] According to various embodiments, electrophoresis (314) is theprocess of providing the samples in the capillaries with mobility by theaction of an electric field created between the cathode and anodebuffers, so that the samples can be separated by differences in mobilitydepending on their characteristics. The separated samples can beoptically detected sequentially in the order of their arrival at thedetector. For example, when the samples are DNA, differences in mobilitycan be caused by their individual base lengths, so that DNA with ashorter base length can move faster and can pass the detector first. Thedetection of DNA by the detector is made possible by attaching afluorescent dye to it. Normally, the measurement time and thevoltage-applied time can be set to accommodate a sample with the longestmigration time.

[0048] According to various embodiments, once a predetermined time haspassed after the application of voltage, and desired data has beenobtained, the application of voltage can be terminated to bringelectrophoresis to an end (315).

[0049] In each of the above-described sequences of preliminary migration(308), sample introduction (311), and electrophoresis (314), a processof detecting abnormalities in the electrophoresis path by monitoringelectric current fluctuations can be carried out. In these threesequences, a high voltage of about 2 kilovolts or more can be applied tothe electrophoresis path. If, for example, the application of a highvoltage is maintained while micro-sized air bubbles (that cannot bedetected by the earlier condition confirmation by a weak voltage) remainin the electrophoresis path, the air bubbles can grow as the temperatureinside the flow path rises. When the current-carrying path is blocked bythe expanding air bubbles, an electric discharge can be caused. For atleast this reason, an apparatus can be equipped with the ability todetect abnormalities in the electrophoresis path while a voltage isapplied.

[0050]FIG. 4 shows a flowchart for a process of detecting abnormalitiesin the electrophoresis path. A method of detecting abnormalities in theelectrophoresis path by monitoring current value fluctuations will bedescribed by referring to FIG. 4.

[0051] First, a voltage can be applied, and the sequence of eitherpreliminary migration, sample introduction, or electrophoresis can beinitiated (401). This can be followed by measuring abnormal currents,and, if an abnormality is detected in the electrophoresis path, theabnormal current measuring function can be placed into operation (402).The value of electric current can then be obtained from the secondammeter 207 (403). If there is a current value that has been acquired inthe past, it can be compared with the latest current value to determinethe condition of the current-carrying path (404). The value of theelectric current is stable value when the current-carrying path isnormal, and any fluctuations are gradual. On the other hand, if theelectrophoresis path is blocked by air bubbles, for example, theelectric current would fluctuate suddenly. Accordingly, by monitoringthe degree of fluctuation in the electric current, and specifically, thegradient of the electric current, abnormalities in the electrophoresispath can be instantly detected while the voltage is applied. Adetermination that there is an abnormality in the current-carrying pathcan be made when the amount of decrease in the current value per unittime has exceeded the threshold. This decision technique will bedescribed in detail below.

[0052] According to various embodiments, when an abnormality is detectedin the current-carrying path, the voltage can be terminated and themeasurement can be stopped to prevent electric discharge. An error canthen be displayed (407), and a response by the operator can be demanded(408). The operator, recognizing the error, can then visually inspectthe electrophoresis path for any air bubbles or foreign substances. Ifan abnormality is found, a countermeasure can be taken, and either theanalysis can be resumed or maintenance by service personnel can berequested. For example, if the cause of the abnormality is air bubblesor the mixing of a foreign substance, the flow path can be refreshed bythe pump device 102, or by manually operating the syringe 106. If noabnormality is found at step (404), either the application of voltagecan be maintained or confirmed (405). When the voltage is to be cut off,the monitoring of the electric current fluctuations can be stoppedbefore cutting off. If the application of voltage is to be continued,the sequence from the electric current value reading step (403) to step(405) can be repeated.

[0053]FIG. 5 schematically illustrates a method of detecting andcomparing electric current fluctuations, and corresponds to the “COMPAREWITH A PREVIOUS VALUE” step of FIG. 4. The method of judging thecondition of the migration path based on an average value of theelectric current will be described by referring to FIG. 5.

[0054] While the obtained electric current values may be compared asthey are in step (404), an average can be taken over a certain timeperiod and then compared, according to various embodiments. Thisoperation has the effect of smoothing the electric current values over acertain period, so that minute fluctuations in the electric currentvalues due to detection accuracy or the influence of sporadicallygenerated static noise can be reduced.

[0055] According to various embodiments, while voltage is being applied,the microprocessor 201 can read the electric current value fluctuationsat periods of about 100 msec, and can check them. The electric currentvalues that are read can be sequentially recorded while they aredesignated as I₁, I₂, I₃, . . . and so on, up to I₁₀, in chronologicalorder. Thereafter, the latest five values, namely I₆ to I₁₀, can beaveraged. The previous five values, namely I₁₁ to I₅, can also beaveraged. As a result, each of the resultant average values can indicatethe average value for a 500-msec interval when the electric currentvalues have been read at 100 msec periods. The difference between theaverage value (I_(ave).(n)) in the latest 500-msec interval and theaverage value (I_(ave).(n−1)) of the previous 500-msec interval can bedefined as a fluctuation (?I) of the electric current value (I).

[0056] In a normal state, the electric current value (I) can assume astable value and, therefore, its fluctuation ?I can be almost zero.However, if the air bubbles in the flow path grow due to the heatproduced by the electric current and they block the flow path, asmentioned above, the electric current value (I) can suddenly decreaseand its fluctuation value ?I can rise. As a result, a certain thresholdcan be set, and the voltage can be removed from the electrophoresis pathimmediately after the electric current value fluctuation has exceededthe threshold. The threshold can be of such a magnitude that it is notobserved in a normal state but can be easily exceeded when anabnormality, such as the blocking of the flow path by air bubbles, isproduced in the electrophoresis path. This kind of desirable thresholdcan be set, for the fluctuation (?I) of the electric current value dueto detection accuracy in a normal state is far smaller than that duringthe occurrence of an abnormality. While the difference between I(n) andI(n−1) can be defined as the fluctuation of the electric current value,the fluctuation of the electric current value can be determined based onthe ratio of I(n) and I(n−1).

[0057] According to various embodiments, it is also possible to detectabnormalities in the electrophoresis path based on the variation of theelectric current value. For example, a threshold can be establishedbased on an expected value of the electric current in the event of thegrowth of air bubbles in the electrophoresis path. The electric currentvalue can be continually compared with the threshold, andelectrophoresis can be terminated if the electric current value dropsbelow the threshold, thereby indicating that the flow path is blocked byair bubbles.

[0058]FIG. 6 shows electric current fluctuation and its detection,showing functional changes from the start of measurement to thedetection of an abnormality. Hereafter, the changes in the actualcurrent value (I) and its fluctuation (?I) will be described. The top ofFIG. 6 shows the time elapsed on the horizontal axis, and the electriccurrent value on the vertical axis. FIG. 6 plots changes in electriccurrent as a voltage is applied at a point and the electric current canthen enter a stable state. This can be followed by the expansion of theair bubbles as the temperature in the flow path increases and the airbubbles blocking the flow path at another point. The bottom of FIG. 6shows the time elapsed on the horizontal axis and the fluctuation of theelectric current as obtained by the method described by referring toFIG. 5. The bottom of the figure shows the fluctuation of the currentvalue from the time when the current value has entered the steady state,until the time when the flow path is blocked by air bubbles.

[0059] As shown in the top of FIG. 6, when a voltage is applied, anelectric current can flow that is commensurate with the applied voltage.In operation, the electric current can be stabilized not long after theapplication of a voltage if there are no abnormalities in theelectrophoresis path. The electric current value (I) can be stabilizedeven if minute air bubbles are mixed in the electrophoresis path if theydo not block the flow path. As the air bubbles grow in size due toheating by applying an electric current great enough such that the airbubbles block the electrophoresis path, the electric current value (I)can greatly decrease. Such a drastic reduction in electric current canalso be caused by an accident, such as short-circuiting in thehigh-voltage source, or by breakage in the electrophoresis path by aninsulator.

[0060] According to various embodiments, the electric currentfluctuation caused by the above electric current changes is shown at thebottom of FIG. 6. While the electric current value (I) is in a stableperiod, the fluctuation (?I) can be nearly zero. Once the electriccurrent value (I) drops due to the growth of air bubbles, the electriccurrent fluctuation (?I) can increase and when it reaches the setthreshold, an abnormal fluctuation in electric current value can bedetected.

[0061] In a system where normally an electric current of several hundredamperes flows, abnormalities in the electrophoresis path, such as thegrowth of air bubbles, can be detected by the above-described method bysetting the threshold at several microamperes.

[0062] According to various embodiments, when the voltage value ischanged during the application of voltage, the electric current cansimultaneously change, and this could result in operational failure.Specifically, the function could make a false detection. In order toprevent this, a method can be employed whereby the abnormal electriccurrent detection function is terminated before the voltage is changed,and then resumed once the electric current has entered a stable region.

[0063] According to various embodiments, another method can be employedwhen the voltage is changed in an increasing direction. Specifically,electric current fluctuation in an increasing direction can be ignoredand fluctuations in a decreasing direction can be monitored, based onthe fact that electric current decreases when air bubbles are produced.In this case, even if the timing of the application of voltage and thatof the detection of abnormal fluctuation are reversed ? that is, even ifthe voltage is applied after the detection of abnormal fluctuation ?there is no possibility of false detection.

[0064] The method of detecting abnormalities in the electrophoresis pathbased on values of electric current and its fluctuation, according tovarious embodiments, can be applied to systems that are susceptible todamage by abnormalities, such as air bubbles, in the electrophoresispath. These include systems that employ a non-fluid separating medium,or systems in which the separating medium in the capillaries is notreplaced for each analysis.

[0065] Those skilled in the art can appreciate from the foregoingdescription that the present teachings can be implemented in a varietyof forms. Therefore, while these teachings have been described inconnection with particular embodiments and examples thereof, the truescope of the present teachings should not be so limited. Various changesand modifications may be made without departing from the scope of theteachings herein.

What is claimed is:
 1. An electrophoresis apparatus comprising: at leastone capillary being filled with a separating medium capable ofseparating a sample, each capillary including an injection end portionfor injection of the sample and a light-illuminable portion capable ofbeing illuminated with light; an excitation optics device capable ofilluminating the light-illuminable portion with light; a light-receivingoptical device capable of detecting light emitted from thelight-illuminable portion; a detection device capable of acquiringinformation from the sample as a function of an electric current flowingin a current-carrying path of the electrophoresis apparatus; and a powersupply device capable of applying a voltage to the current-carrying pathincluding at least the injection end portion and the light-illuminableportion of each capillary, and capable of adjusting or terminating thevoltage based on the information acquired by the detection device. 2.The electrophoresis apparatus of claim 1, wherein the power supplydevice is capable of adjusting or terminating the voltage based on arate of change of the electric current.
 3. The electrophoresis apparatusof claim 2, wherein the power supply device is capable of adjusting orterminating the voltage when the rate of change of the electric currenthas exceeded a predetermined value.
 4. The electrophoresis apparatus ofclaim 3, wherein the predetermined value of the electric current can beset depending on at least one of a length of the capillary, the numberof the capillaries, the separating medium, and the method ofmeasurement.
 5. The electrophoresis apparatus of claim 1, wherein thepower supply device is capable of adjusting or terminating the voltagewhen the current has reached a predetermined value.
 6. Theelectrophoresis apparatus of claim 1, wherein the power supply device iscapable of continuing to supply the voltage based on the informationacquired by the detection device when the voltage is changed.
 7. Theelectrophoresis apparatus of claim 1, wherein the power supply device iscapable of adjusting or terminating the voltage when air bubbles areproduced in the separating medium.
 8. The electrophoresis apparatus ofclaim 1, further comprising an information-transmitting device that iscapable of indicating that air bubbles have formed in the separatingmedium based on the information acquired by the detection device.
 9. Amethod of performing electrophoresis comprising: providing a capillaryincluding an injection end portion for injection of a sample and alight-illuminable portion capable of being illuminated with light;detecting a value of an electric current flowing in a separating mediumarranged in the capillary while performing electrophoresis; andterminating performing electrophoresis based on the detected value ofthe electric current.
 10. The method of claim 9, wherein electrophoresisis terminated when a rate of change of the detected value of theelectric current has exceeded a predetermined value.
 11. The method ofclaim 9, wherein detecting a value of an electric current includescalculating an average value of the electric current over apredetermined period of time, and electrophoresis is terminated when therate of change of the average value of the electric current exceeds apredetermined value.
 12. The method of claim 9, wherein electrophoresisis terminated when the detected value of the electric current dropsbelow a predetermined value.
 13. The method of claim 10, wherein thepredetermined value is determined based on at least one of a length of acapillary, the number of capillaries, the separating medium, and themethod of measurement.
 14. An electrophoresis apparatus comprising: atleast one capillary being filled with a separating medium capable ofseparating a sample, each capillary including an injection end portionfor injection of the sample and a light-illuminable portion capable ofbeing illuminated with light; an excitation optics device capable ofilluminating the light-illuminable portion with light; a light-receivingoptical device capable of detecting light emitted from thelight-illuminable portion; a detection device capable of acquiringinformation from the sample as a function of an electric current flowingin a current-carrying path of the electrophoresis apparatus; a powersupply device capable of applying a predetermined voltage to thecurrent-carrying path including at least the injection end portion andthe light-illuminable portion of each capillary; and an indicator devicecapable of indicating the state of the current-carrying path based onthe current flowing in the current-carrying path upon applying thepredetermined voltage.
 15. The electrophoresis apparatus of claim 14,wherein the indicator device is capable of indicating that a conditionof the current-carrying path is not suitable for electrophoresis whenthe current is smaller than a predetermined value.
 16. Theelectrophoresis apparatus of claim 14, wherein the predetermined voltageis smaller than a voltage that is capable of producing an electricdischarge when the at least one capillary is not filled with theseparating medium.
 17. The electrophoresis apparatus of claim 14,further comprising a buffer-solution container carrying a buffersolution that constitutes part of the current-carrying path, wherein thepredetermined voltage is smaller than a voltage that is capable ofproducing an electric discharge when the buffer-solution container isnot filled with the buffer solution.
 18. A method of performingelectrophoresis utilizing a capillary, the method comprising: fillingthe capillary with a separating medium; applying a voltage to acurrent-carrying path including the separating medium prior toperforming electrophoresis, the applied voltage being smaller than avoltage that is applied during electrophoresis; detecting a first valueof the electric current flowing in the current-carrying path; anddetermining a state of the current-carrying path based on the firstvalue of the electric current that is detected.
 19. The electrophoresismethod of claim 18, wherein the state that the current-carrying path isnot suitable for electrophoresis is determined when the first value ofthe electric current that is detected is smaller than a predeterminedvalue.
 20. The electrophoresis method of claim 18, wherein the voltagethat is applied to the current-carrying path is smaller than a voltagethat produces electric discharge when the capillary is not filled withthe separating medium, or when a buffer-solution container forming partof the current-carrying path does not contain an appropriate amount of abuffer solution.
 21. The electrophoresis method of claim 18, furthercomprising performing electrophoresis while detecting a second value ofthe electric current flowing in the separating medium after determiningthat a state of the current-carrying path is suitable forelectrophoresis, and terminating performing electrophoresis based on thedetected second value of the electric current.
 22. The electrophoresismethod of claim 21, wherein electrophoresis is terminated when a rate ofchange of the detected second value of the electric current has exceededa predetermined value.
 23. The electrophoresis method of claim 21,wherein detecting a second value of an electric current includescalculating an average value of the electric current over apredetermined period of time, and electrophoresis is terminated when therate of change of the average value of the electric current exceeds apredetermined value.
 24. The electrophoresis method of claim 21, whereinelectrophoresis is terminated when the detected second value of theelectric current drops below a predetermined value.
 25. Theelectrophoresis method of claim 22, wherein the predetermined value isdetermined based on at least one of a length of a capillary, the numberof capillaries, the separating medium, and the method of measurement.26. An electrophoresis apparatus comprising: at least one capillarycapable of being filled with a separating medium capable of separating asample, each capillary including an injection end portion for injectionof the sample and a light-illuminable portion capable of beingilluminated with light; an excitation optics device capable ofilluminating the light-illuminable portion with light; a light-receivingoptical device capable of detecting light emitted from thelight-illuminable portion; a detection device capable of acquiringinformation from the sample as a function of an electric current flowingin a current-carrying path of the electrophoresis apparatus; and a powersupply device capable of applying a voltage to the current-carrying pathincluding at least the injection end portion and the light-illuminableportion of each capillary, and capable of adjusting or terminating thevoltage based on the information acquired by the detection device. 27.The electrophoresis apparatus of claim 26, wherein the power supplydevice is capable of adjusting or terminating the voltage based on arate of change of the electric current.
 28. The electrophoresisapparatus of claim 27, wherein the power supply device is capable ofadjusting or terminating the voltage when the rate of change of theelectric current has exceeded a predetermined value.
 29. Theelectrophoresis apparatus of claim 28, wherein the predetermined valueof the electric current can be set depending on at least one of a lengthof the capillary, the number of the capillaries, the separating medium,and the method of measurement.
 30. The electrophoresis apparatus ofclaim 26, wherein the power supply device is capable of adjusting orterminating the voltage when the current has reached a predeterminedvalue.
 31. The electrophoresis apparatus of claim 26, wherein the powersupply device is capable of continuing to supply the voltage based onthe information acquired by the detection device when the voltage ischanged.
 32. The electrophoresis apparatus of claim 26, wherein thepower supply device is capable of adjusting or terminating the voltagewhen air bubbles are produced in the separating medium.
 33. Theelectrophoresis apparatus of claim 26, further comprising aninformation-transmitting device that is capable of indicating that airbubbles have formed in the separating medium based on the informationacquired by the detection device.
 34. An electrophoresis apparatuscomprising: at least one capillary capable of being filled with aseparating medium capable of separating a sample, each capillaryincluding an injection end portion for injection of the sample and alight-illuminable portion capable of being illuminated with light; anexcitation optics device capable of illuminating the light-illuminableportion with light; a light-receiving optical device capable ofdetecting light emitted from the light-illuminable portion; a detectiondevice capable of acquiring information from the sample as a function ofan electric current flowing in a current-carrying path of theelectrophoresis apparatus; a power supply device capable of applying apredetermined voltage to the current-carrying path including at leastthe injection end portion and the light-illuminable portion of eachcapillary; and an indicator device capable of indicating the state ofthe current-carrying path based on the current flowing in thecurrent-carrying path upon applying the predetermined voltage.
 35. Theelectrophoresis apparatus of claim 34, wherein the indicator device iscapable of indicating that a condition of the current-carrying path isnot suitable for electrophoresis when the current is smaller than apredetermined value.
 36. The electrophoresis apparatus of claim 34,wherein the predetermined voltage is smaller than a voltage that iscapable of producing an electric discharge when the at least onecapillary is not filled with the separating medium.
 37. Theelectrophoresis apparatus of claim 34, further comprising abuffer-solution container carrying a buffer solution that constitutespart of the current-carrying path, wherein the predetermined voltage issmaller than a voltage that is capable of producing an electricdischarge when the buffer-solution container is not filled with thebuffer solution.